Project Summary Protein arginine methyltransferase 5 (PRMT5) is part of a methyltransferase complex that symmetrically dimethylates arginine residues in a wide range of proteins responsible for many biological processes. PRMT5 inhibitors (PRMT5i) have been developed and are currently in clinical trials. However, the determinants of PRMT5i response and resistance are largely unknown. I will elucidate the molecular mechanisms of PRMT5i response and resistance in two tumor types, lung adenocarcinoma (LUAD) and pancreatic ductal adenocarcinoma (PDAC). Our lab has identified multiple sensitive parental (P) LUAD and PDAC lines and generated independent resistant (R) variants. Experiments in LUAD show that the R cells all arise through a single mechanism, which is drug-induced, not pre-existing, and reflects a dramatic shift in gene expression. I hypothesize that this new gene expression signature enables the R cells to tolerate PRMT5 inhibition and avoid proliferation defects. We have already identified one gene, Stmn2, as being essential to maintain PRMT5i resistance. In Aim 1, I will dissect how STMN2 mediates this effect, and will also identify and characterize additional regulators of resistance through CRISPR/Cas9 KO screens. I hypothesize that adoption of an alternate cell state will be the common mechanism by which tumors escape PRMT5 inhibition, irrespective of their tissue type. In Aim 2, I will test this idea by interrogating the mechanisms of PRMT5i resistance in a second tumor type, PDAC, bearing the same K-rasG12D and tp53 driver mutations as the LUAD cells. Overall, results from this project will provide key insights into the core biological processes that enable PRMT5i sensitivity and resistance, and establish the degree to which these are conserved between, or unique to, different tumor types. This will greatly advance the mechanistic understanding of PRMT5i therapies as these drugs are entering the clinic. While on this fellowship, approximately 70-80% of my effort will be focused on research related activities, including reading literature, doing benchwork, interacting with my PI, lab teams and collaborators to discuss my (and their) work, and presenting in lab meetings. The remaining 20-30% of my effort will be focused on activities that will enable my career development, including writing, oral speaking, and mentoring. Importantly, the community and resources available in the Koch Institute for Integrative Cancer Research at MIT provides an exceptional scientific and intellectual environment to advance my scientific independence.